Ectoparasiticidal aqueous suspension formulations of spinosyns

ABSTRACT

The invention provides a stable ectoparasiticidal aqueous suspension formulation of a spinosyn, comprising the spinosyn, or a physiologically acceptable derivative or salt thereof, milled to an average particle size of 1 to 15 microns, a surfactant in an amount effective to facilitate wetting the milled particles; a dispersant in an amount that forms a spinosyn;dispersant weight ration of from 3:1 to 1:5; and water. It also provides a method of controlling an ectoparasite infestation on a small ruminant or companion animal comprising administering an effective amount of such an aqueous suspension formulation.

This application is the National Stage of International Application No.PCT/US00/19558, filed Aug. 2, 2000, which claims the benefit of U.S.Provisional Application Ser. No. 60/148,527, filed Aug. 12, 1999.

There are many types of ectoparasiticidal formulations. These typesinclude emulsifiable concentrates, wettable powders, organic solventsolutions and suspensions. Many of these formulations require the use ofan organic solvent. For example, an organic solvent must be used whenpreparing an oil-in-water emulsifiable concentrate. Organic solvents,however, are typically regarded as having adverse environmental orecological effects, and they can add to the overall toxicity of theformulation. Wettable powders can be dispersed in tank mix formulationswithout organic solvents, but they are generally inferior to otherformulations in biological effect and handling characteristics. There isa need, therefore, for safer formulations such as aqueous formulations.

Spinosyns (also known as A83453 factors) are known agriculturalinsecticides. Because of their low toxicity to animals and humans,spinosyns are considered to be environment-friendly, “green” pesticides.It is desirable to formulate spinosyns to maintain this “green” profile.

The spinosyns were also known to have some ectoparasiticidal activity,i.e., they had in vitro activity against mosquito larvae, black blowflylarvae and adult stable flies, which are members of the insect orderDiptera, and transient systemic activity against larval blowfly andadult stable fly in guinea pigs and sheep. For these studies, thespinosyns were administered in aqueous polyvinylpyrrolidone or inpolyethylene glycol (see, U.S. Pat. No. 5,571,901, col. 26-32).

The spinosyns have recently been found to be useful in controllingectoparasites on sheep and companion animals. Thus, useful formulationsof spinosyns with low toxicity and increased stability are potentiallyvaluable in combating ectoparasites, thereby preventing the diseasessuch pests often carry.

Aqueous formulations of spinosyns would be most desirable.Unfortunately, spinosyns have low solubility in water and are unstablein aqueous solution.

This invention provides a stable aqueous suspension formulation suitablefor spinosyns. These aqueous suspension formulations offer severaladvantages over previous non-aqueous or solvent-containing spinosynformulations. Their advantages include greater chemical, biological andthermal stability and improved ease of use.

The ratio of active ingredient to dispersant is a unique characteristicof the formulations of this invention. Generally, aqueous suspensionformulations have a ratio of active ingredient to dispersant ratio inthe range of about 5:1 to about 25:1. The formulations of thisinvention, however, have higher amounts of dispersant, bringing thespinosyn to dispersant ratio to from about 3:1 to about 1:5. Previousformulations of spinosyns with relatively low concentrations ofdispersant, as compared to the higher concentrations in the presentformulations, tended to lack homogeneity and predictability with respectto expected concentrations upon dilution. This result was surprisingbecause it was thought that spinosyns at such low concentrations wouldbe completely solubilized.

Increasing the concentration of dispersant in the aqueous suspensionsproduced another unexpected result. When the formulations with increaseddispersant concentration were diluted to form aqueous dip solutionscontaining 5 ppm to 25 ppm of spinosyn, the diluted solutions hadhomogenous spinosyn concentrations throughout the dip solution, a verybeneficial effect.

In particular, this invention provides a stable ectoparasiticidalaqueous suspension formulation comprising an ectoparasiticidal amount ofa spinosyn, or a physiologically acceptable derivative or salt thereof,milled to an average particle size of about 1 to about 15 microns, and asurfactant in an amount effective to facilitate wetting the milledparticles; a disperant in an amount sufficient to form aspinosyn:dispersant weight ratio of from 3:1 to about 1:5; and water.

Particularly useful stable ectoparasiticidal aqueous suspensionformulations of this invention are those;

a) wherein the amount of spinosyn is from about 0.1 to about 50 weightpercent of the formulation;

b) wherein the dispersant is ionic;

c) wherein the amount of surfactant is about 0.1 to about 10 weightpercent of the formulation;

d) which further comprise about 0.3 to about 5 weight percent of amineral thickener,

e) which further comprise about 0.05 to about 3 weight percent of a gum,and

f) which further comprise an antimicrobial agent acceptable for topicalveterinary applications in an amount effective to prevent microbialgrowth in the suspension.

Other preferred spinosyn-containing formulations comprise about 25gram/liter of spinosad, a condensed napthalene sulfonic acid as adispersant, propylene glycol as an antifreeze agent and humectant, asurfactant, a mineral suspending aid, a xanthan gum suspending aid, anantimicrobial agent, a foam control agent, and deionized water(vehicle).

The components can be mixed in various proportions to achieve thecharacteristics desired in the formulation.

The formulations of this invention are aqueous suspensions. By “aqueous”is meant that the formulation is a water-based system, i.e., no organicsolvents are included in the formulation.

The fact that the present compositions are water-based is important froma chemical stability perspective. Aqueous suspension formulations ofthis invention containing 25 g/L of spinosad have been shown to bechemically stable at ambient and elevated temperatures for at least sixmonths as indicated by HPLC analysis. The formulations are physicallystable and readily dispersible in water for use. For topical dips,sprays, and other applications, having the spinosyn delivered in wateris a great advantage. The formulations can be used without dilution(neat), either as a pour-on or spot-on, or they can be diluted to forman homogeneous aqueous solution suitable for use as a topical dip.

An unexpected advantage is that these formulations provide whole animalectoparasiticidal effectiveness when applied as a pour-on or spot-onapplication. When the formulations are used as topical dips, forexample, they allow easy whole treatment of larger animals such assheep, goats, and camellids, etc., with minimal “stripping” of theformulation from the diluted dip as the number of animals treated in adip pool increases.

The insecticidal component in these formulations is a spinosyn, or aderivative or salt thereof. Spinosyns are naturally-derived macrolidesproduced by fermentation of Saccharopolyspora spinosa. The fermentationproduces multiple factors, including spinosyn A and spinosyn D (alsocalled A83543A and A8354D). Spinosyn A and spinosyn D are the twospinosyns that are most active as insecticides. An agricultural productcomprised mainly of these two spinosyns is available commercially underthe generic name “spinosad” for field applications.

Spinosyn A was the first spinosyn isolated and identified from thefermentation broth of Saccharopolyspora spinosa. Subsequent examinationof the fermentation broth revealed that S. spinosa produced a number ofspinosyns that have been called spinosyns A to J (A83543A to J).Additional spinosyns, denominated K to W, have been identified frommutant strains of S. spinosa. The various spinosyns are characterized bydifferences in the substitution patterns on the amino group of theforosamine, at selected sites on the tetracyclic ring system and on the2N,3N,4N-(tri-O-methyl)rhamnose group.

The term “spinosyn” as used herein refers to one or more spinosyn factor(spinosyn A, B, C, D, E, F, G, H, J, K, L, M, N, O, P, Q, R, S, T, U, V,W or Y), an N-methyl derivative of one or more spinosyn factor, or acombination thereof. For convenience, the term “spinosyn” or “spinosyncomponent” will also be used herein to mean a spinosyn factor, aphysiologically acceptable derivative or salt of a spinosyn factor, or acombination thereof.

Boeck et al. described spinosyns A-H and J (which they called A83543factors A, B, C, D, E, F, G, H and J), and salts thereof in U.S. Pat.No. 5,362,634 (issued Nov. 8, 1994); U.S. Pat. No. 5,496,932 (issuedMar. 5, 1996); and U.S. Pat. No. 5,571,901 (issued Nov. 5, 1996).Mynderse et al. described spinosyns L-N (which they called A83543factors L, M and N), their N-dimethyl derivatives, and salts thereof, inU.S. Pat. No. 5,202,242 (issued Apr. 13, 1993); and Turner et al.described spinosyns Q-T (which they called A83543 factors Q, R, S andT), their N-dimethyl derivatives, and salts thereof, in U.S. Pat. No.5,591,606 (issued Jan. 7, 1997) and U.S. Pat. No. 5,631,155 (issued May29, 1997). Spinosyns K, O, P, U, V, W and Y are described, for example,by Carl V. DeAmicis, James E. Dripps, Chris J. Hatton and Laura I. Karrin American Chemical Society's Symposium Series: Phytochemicals for PestControl, Chapter 11, “Physical and Biological Properties of Spinosyns:Novel Macrolide Pest-Control Agents from Fermentation”, pages 146-154(1997).

The spinosyns can be isolated in the form of salts that are also usefulin the compositions and methods of this invention. The salts areprepared using standard procedures for salt preparation. For example,spinosyn A can be neutralized with an appropriate acid to form an acidaddition salt. Representative suitable acid addition salts include saltsformed by reaction with either an organic or inorganic acid, forexample, sulfuric, hydrochloric, phosphoric, acetic, succinic, citric,lactic, maleic, fumaric, cholic, pamoic, mucic, glutamic, camphoric,glutaric, glycolic, phthalic, tartaric, formic, lauric, stearic,salicylic, methanesulfonic, benzenesulfonic, sorbic, picric, benzoic,cinnamic and like acids.

When preparing the formulations of this invention, the spinosyncomponent should be milled to an average particle size of from about 1to about 15 microns in order to form the most suitable suspension. Apreferred average particle size is from about 2 to about 7 microns,especially 3 to 7 microns. The milling is accomplished by a “wetmilling” process in which the spinosyn is exposed to sufficientsurfactant to facilitate wetting the milled particles.

The formulations of this invention comprise an ectoparasiticidal amountof the spinosyn component. By “ectoparasiticidal amount” is meant anamount that effectively controls or kills a target insect, parasite, orectoparasite when applied to an animal that either has an insect,parasite or ectoparasite infestation or is susceptible to acquiring suchan infestation. As those in the art understand, the amount of spinosynthat is ectoparasiticidal will vary, depending upon a number of factors,including the insect or parasite being controlled, the host animal beingtreated, other components of the formulation and the route ofadministration.

In order to provide an ectoparasiticidal amount and form a suitablesuspension, the spinosyn concentration should be in the range of fromabout 0.02 to about 50 percent by weight of the formulation. Preferably,the spinosyn concentration should be in a range of from about 0.1 toabout 50 weight percent of the formulation Even more preferably, thespinosyn component is present in an amount of from about 2 to about 5weight percent of the formulation. For example, a useful formulation isone wherein 25 g of the spinosyn component is present per liter of theformulation.

The formulations of this invention also include a dispersant. Adispersant is a compound that is able to counteract particle-to-particleattraction within an aqueous suspension without significant reduction insurface tension of the aqueous suspension vehicle (i.e., the addition ofthe dispersant does not reduce the surface tension of water below 40dynes/cm). A dispersant has physicochemical properties that allow thedispersant to orient itself between particles of active ingredient and,by virtue of the dispersant's size and/or charge, reduce thecohesiveness or attraction of the active ingredient particles for eachother. In addition to imparting physical stability to the aqueousmixture, dispersants may also aid in the redispersibility of a dilutedspray mixture. The dispersing agent should be carefully selected andused to avoid problems such as agglomeration, sedimentation andflocculation. Any dispersant that interferes with particle-to-particleattraction or cohesiveness by virtue of size and/or charge is useful asa dispersant for purposes of the present invention.

Both ionic and nonionic dispersants are useful in the formulations ofthis invention, but ionic dispersants are preferred. Examples includelignosulfonic acids and salts thereof, polymerized alkyl, arylalkyl ornaphthalene sulfonic salts, comb polymeric dispersants (such as ATLOX4913™, Uniqema), condensed formaldehyde/naphthalene sulfonic acid andsalts thereof, sodium dioctyl sulfosuccinate and high molecular weightanionic dispersants. An especially useful dispersant is a condensedformaldehyde/napthalene sulfonic acid or a salt thereof. A suitablecondensed napthalene sulfonic acid dispersant is available from KenkelCorp. as LOMAR PWA.

The type of water used in these aqueous formulations is not critical.For example, it can be tap water or deionized water. The water can havea pH range of from about 5 to about 10, with an ideal range pH of 6 to9.

The surfactant used in the formulations of this invention shouldmaintain the resulting suspension of milled particles at a low viscosityand allow a high percentage recovery of milled solids after processing.A surfactant is a compound that is surface active and reduces thesurface tension of water to ≦40 dynes/cm. Although anionic, cationic,nonionic and amphoteric surfactants can be used in these formulations,nonionic surfactants are preferred. Examples of surfactants that areparticularly useful include straight and branched chain octyl and nonylphenols, straight and branched chain alcohol ethoxylates, and alkyl arylether ethoxylates.

The surfactant should be present in an amount sufficient to facilitatewetting the milled particles of the spinosyn component. Generally, theamount of surfactant is from about 0.1 to about 10 weight percent of theformulation. A preferred amount of surfactant is from about 0.1 to about5 weight percent of the formulation.

Often nonionic surfactants will efficiently wet solids without tendingto solubilize micron-sized particles after milling. Certain blockcopolymers of polyoxypropylene-polyoxyethylene that contain ethyleneoxide are particularly useful surfactants in the formulations. Thesesurfactants vary in wetting ability as the ethylene oxide contentvaries. Examples are the PLURONIC series (BASF). PLURONIC P-103™,PLURONIC P-104™, and PLURONIC P-123™ surfactants are especiallypreferred. Qualitative wetting tests of these surfactants in waterindicated an ability to wet technical grade spinosad in less than 30seconds.

A number of other optional components may be added to the formulationsof this invention. Examples of these include:

-   suspending aids or thickeners, UV absorbing compounds,-   antimicrobial agents, viscosity modifying compounds,-   antifoam agents or defoamers, dyes,-   substantive agents, perfumes,-   antifreeze agent, deodorants,-   humectants, and-   physiologically or dermatologically acceptable carriers, diluents,    excipients or adjuvants.

Suspending aids or thickeners aid in structure formation and rheologybuilding of the aqueous suspension formulations. These agents impartphysical stability to the suspensions. Thickeners increase the viscosityof the formulation, thereby aiding in the suspension of activeingredient. Many types of thickeners are available. These include gumsand natural polysaccharides, mineral thickeners, and synthetic polymericthickeners.

The gums and natural polysaccharides class of thickeners includesnumerous gums, starches, celluloses, and other polysaccharides. Examplesof gums and natural polysaccharides are xanthan gum, guar gum, locustbean gum, carrageenan, pectin, tragacanth and tamarind gum.

Examples of mineral thickeners are inorganic clays, fumed andprecipitated silica, mixed metal hydroxides and mixed metal silicates.Among the inorganic thickeners are various commercially available silicathickeners, including hydrophilic silicas and hydrophobic silicas.Hydrophobic amorphous fumed silicas are also useful as the thickeningadditive. Examples of hydrophobic silicas are AEROSIL R-972 and AEROSILR-974 from Degussa Corporation, Akron, Ohio.

A preferred mineral thickener for use in the formulations is a complexcolloidal magnesium aluminum silicate refined and derived from naturalsmectite clays. R. T. Vanderbilt Co. makes a suitable mineral suspendingaid called VEEGUM and a xanthan gum suspending aid called RHODOPOL 23.

Synthetic polymeric thickeners arc anionic, nonionic, cationic orhydrophobically modified polymers. Examples include compounds such assodium polyacrylates, alkyl and alkyloxycelluloses (including sodiumcarboxymethyl cellulose, methyl cellulose, ethoxylated cellulose,hydroxypropylmethyl cellulose, hydroxyethyl cellulose and modifiedhydroxyethyl cellulose), microcrystalline cellulose, starches andmodified starches, polyvinylpyrrolidone, polyethylene glycol ofmolecular weight from 2000 to 4,000,000, and mixtures thereof.Preferably, the polymer is selected from the group consisting of sodiumpolyacrylate, hydroxyethyl cellulose, cetyl hydroxyethyl cellulose,polyvinylpyrrolidone and polyquaternium-10.

Other compounds are also useful as thickening agents or suspending aidsin the formulations. For example, sugars, salts and other smallmolecules such as urea can be used to increase the density of the waterused in the aqueous formulation, thus aiding in the suspension of theactive ingredient particles. These compounds are added to theformulation in an amount sufficient to increase the density of theaqueous solution to counteract the physical forces that favor settlingout of the suspension particles.

The amount of thickener or suspending aid and the ratio of thesuspending aid to the spinosyn component vary depending on the desiredconcentration of the spinosyn component in the formulation. In general,the amount of thickener is from about 0.05 to about 8 weight percent ofthe formulation. For example, a useful formulation containing 200 g/L ofspinosad and 0.2% (w/w) xanthan gum contained 1% (w/w) of complexcolloidal magnesium aluminum silicate (1:5 ratio), whereas formulationswith lower amounts of active or solids content, such as 50 g/L or 25 g/Lof spinosad, and 0.2% (w/w) xanthan gum contained 2% (w/w) of complexcolloidal magnesium aluminum silicate (1;10 ratio) in the finalformulation.

As a general rule, suspensions with a higher content of solids or activeingredient are more efficient to wet mill and suspend in water thansuspensions with lower content of solids or active ingredient.Suspensions with a lower concentration of active ingredient have a highwater content, requiring the addition of proportionately higher amountsof suspending aids to suspend the solids. The viscosity of the finalaqueous suspension formulation should be at a minimum so it can beeasily poured from the container and mixed with water for use.

Antimicrobial agents are often added to formulations to prevent unwantedmicrobial growth, particularly if a component of the formulationsupports such growth. For example, when xanthan gum is used as athickening agent to suspend solids and build viscosity, the addition ofan antimicrobial agent prevents microbial attack of the gum and loss ofproduct viscosity.

A variety of antimicrobial agents are useful for this purpose. Certainchemicals such as 1-(3-chloroallyl)-3,5,7-triaza-1-azoniaadamantanechloride and 1,2-benzisothiazolin-3-one are examples of particularlyuseful antimicrobial agents. The former compound is available from DowChemical Company as DOWICIL 75™ preservative; and an aqueous solution ofthe latter in di(propylene glycol) is available from Zeneca Biocides asPROXEL GXL™. The latter antimicrobial agent is stable in the presence ofamines and amine-containing compounds such as spinosad and possesses abroad spectrum of activity against microorganisms.

When the antimicrobial component is a liquid and the thickening agent issuspended in an agent such as propylene glycol before hydration, it isconvenient to incorporate the antimicrobial agent in the suspendingagent.

Typically, the aqueous flowable formulations of this invention maycontain from about 0.01 to about 0.5 weight percent of an antimicrobialagent, with a preferred range of 0.04 to about 0.3 weight percent. Forexample, a formulation of this invention contained xanthan gum as athickener and an aqueous solution of 1,2-benzisothiazolin-3-one indi(propylene glycol) at a level of 0.2% w/w (2000 ppm). This level ofantimicrobial agent was effective to preserve the gum.

An antifoam agent or defoamer is a useful optional component in thepresent formulations. Poly(dimethylsiloxane) antifoams are particularlyuseful. They can be initially incorporated into the grind batch ofspinosad for foam control. Examples of these agents are ANTIFOAM A™ andANTIFOAM C™, available from Dow Corning. The former is a 100% activeantifoam. The latter, which is a 30% emulsion of poly(dimethylsiloxane)in water, was found to be more effective in processing. It isadvantageously incorporated into the formulation at about 0.2% w/w.Other examples of commercially available antifoam agents useful in thepresent formulations include ANTIFOAM FG-10, ANTIFOAM DB-100, andANTIFOAM AF-100 (all available from Dow Corning). Other antifoam agentsare also useful in these formulations.

A substantive agent is another ingredient that may be added to thepresent formulations. The term “substantive agent” means a compound thatincreases the binding or retention of an active ingredient (in this casethe spinosyn component) to the surface layer of the stratum corneum orto hair. Preferred substantive agents also aid in resisting removal ofthe active component by water or by physical contact, such as rubbing.Examples of useful substantive agents include acrylates, polyvinylacetates, and polyvinyl alcohols.

This invention also relates to a method of manufacturing a stableectoparasiticidal aqueous suspension formulation, said methodcomprising:

(1) wet-milling a composition containing a spinosyn, or aphysiologically acceptable derivative or salt thereof, with asurfactant, a dispersant, an antifoam agent and water to form a “grindcomposition” in which the spinosyn has an average particle size is fromabout 1 to about 15 microns;

(2) blending an aqueous suspension containing about 2 to about 10percent by weight of a mineral thickener with a dispersion compositioncontaining about 1 to about 4 percent by weight of a gum in a C₂-C₄alkylene diol to form a “hydrated suspension composition” containingabout 0.5 to about 8 percent by weight of the mineral thickener; and

(3) diluting a first volume of the grind composition with a secondvolume of the hydrated suspension composition sufficient to provide thedesired spinosyn concentration.

Alternatively, the formulations of this invention can be prepared by (1)making a concentrated aqueous suspension of the spinosyn component, (2)diluting the concentrate to appropriate spinosyn concentrations for useas a pour-on, spot-on or dip concentrate, and then (3) adding asufficient amount of dispersant to bring the ratio of spinosyn todispersant into the range of about 3:1 to about 1:5.

A nonionic surfactant (such as PLURONIC P-123™) is a preferredsurfactant to incorporate into the aqueous suspension of spinosyn. Asuitable “hydrated suspension batch” is formed by blending a hydratedsuspension of a complex colloidal magnesium aluminum silicate suspendingaid (such as VEEGUM) with a xanthan gum hydrated in propylene glycol(such as RHODOPOL 23™). The hydrated suspension batch and additionalwater as needed are blended with the grind batch to prevent syneresis,or separation of clear watery fluid from suspended milled solids. Theappropriate amount of the hydrated suspension batch to be blended withthe grind batch to complete the formulation is determined based upon thepercent recovery of the grind batch after particle size reduction.

The following order of addition of formulation inerts is recommended toprepare the hydrated suspension, which is used to control productviscosity and prevent syneresis of milled spinosad: (1) add all thecomplex colloidal magnesium aluminum silicate to deionized water withhigh speed stirring and allow to fully hydrate; (2) add the xanthan gumto the propylene glycol with stirring to fully disperse the gum in theglycol; and (3) instantly hydrate the xanthan gum in water by theaddition of item (2) to item (1) with stirring. Do not incorporateexcessive amounts of air into the suspension.

In another aspect, this invention provides an article of manufacture,comprising packaging material and a formulation for controlling anectoparasite infestation on a small ruminant or companion animalcontained within said packaging material, wherein said formulationcomprises:

-   -   a) a unit dose of an ectoparasiticidal amount of a spinosyn, or        a physiologically acceptable derivative or salt thereof, milled        to an average particle size of from about 1 to about 15 microns,        and a surfactant in an amount effective to facilitate wetting        the milled particles;    -   b) a dispersant in an amount sufficient to form a        spinosyn:dispersant weight ratio of about 3:1 to about 1:5; and

c) water; and

wherein said packaging material comprises a label or package insert withinstructions for administering the dose to the animal.

This invention also encompasses a method of controlling an ectoparasiteinfestation on a small ruminant or companion animal, comprisingadministering to the animal a formulation comprising anectoparasiticidal amount of a spinosyn, or a physiologically acceptablederivative or salt thereof, milled to an average particle size of fromabout 1 to about 15 microns, and a surfactant in an amount effective tofacilitate wetting the milled particles; a dispersant in an amountsufficient to form a spinosyn:dispersant weight ratio of from 3:1 to1:5; and water.

The term “controlling” refers to either eliminating or ameliorating acurrent infestation or preventing an infestation on a susceptibleanimal. By “animal” is meant a small ruminant or a companion animal.Small ruminants include sheep, goats and camellids. Examples ofcompanion animals are dogs, cats, horses and other pets owned andmaintained in close association with humans as part of the human-animalbond.

A preferred formulation for use in this method is an aqueous suspensioncomprising from about 1 to about 50 weight percent of a spinosyn, adispersant in an amount sufficient to bring the spinosyn:dispersantratio to about 3:1 to about 1:5, and about 0.1 to about 5 weight percentof a surfactant.

In this method, the aqueous suspension is preferably applied topicallyin a pour-on or spot-on treatment protocol. In a pour-on or spot-ontreatment, the formulation is applied directly to the animal's hairand/or skin on the head, neck, shoulders or back, with the treated areabeing less than 10 percent of the surface area of the hair and skin ofthe animal.

Optional ingredients that can be included in the aqueous suspension usedas a pour-on or spot-on include about 1 to about 5 weight percent of asuspending aid selected from mineral thickeners and gums, about 0.5 toabout 2 weight percent of an ionic dispersant, up to 10 percent (w/w) ofa polymeric substantive agent to increase substantiveness of theformulation to hair and/or skin, and an antimicrobial agent in an amounteffective to prevent the growth of microorganisms in the aqueoussuspension.

An advantage of this method, and of the aqueous suspensions of thisinvention, is that the spinosyn only needs to be applied weekly orbi-weekly. This characteristic allows the animal's caretaker to minimizethe effort needed to control the ectoparasites on the animal bylengthening the period between applications. Another advantage is thatthe formulations can be applied rapidly and easily. Further, the cost ofapplication equipment used with these formulations is very low incomparison with that required for other ectoparasiticide formulations.

In addition to the pour-on and spot-on applications, the aqueousspinosyn suspensions of this invention can also be used inwater-dilutable dip and spray applications. Further, the aqueoussuspension formulations can also be useful for systemic administrationof the active ingredient, such as by use in feed or as an injectableformulation.

The following examples illustrate the formulations of this invention. Inthe examples, the spinosad used (“spinosad, technical grade”) was aproduct available commercially from Dow Agrosciences. In preparing theformulations the spinosad was milled to a particle size of from 3 to 7microns.

EXAMPLE 1 Effect of Dispersant on Spinosad Concentration

To examine whether addition of a dispersant allows for greaterpredictability of diluted concentrations when compared to aqueoussuspensions lacking such dispersants, laboratory studies were conductedin which aqueous suspensions were evaluated in the presence and absenceof dispersant. Two groups of aqueous suspensions containing 25 g/L ofspinosad were prepared, one group containing a dispersant and onewithout dispersant. The dispersant used was an ammonium salt ofsulfonated naphthalene condensate that was about 45% solids. It was usedat concentrations of 4 to 5% weight/weight to give about 2% activedispersant on a solids basis.

The suspensions were diluted in a sufficient amount of tap water ordeionized water at various pH levels to dilute the spinosad to atheoretical concentration of 100 ppm. Samples were evaluated for actualspinosad concentration immediately upon dilution and after 24 hours.

Table 1 compares the spinosad concentrations in two types of water atthree pH levels at the time the samples were initially diluted to atheoretical concentration of 100 ppm and 24 hours after dilution.

TABLE 1 Effect of Dispersant on Spinosad Concentration SPINOSAD (ppm) 24Hr. Post-dilution Initial Concentration Concentration Water NoDispersant Dispersant No Dispersant Dispersant Deionized (pH = 4.0) 74.192.7 59.7 70.4 (pH = 7.0) 65.1 92.2 27.8 73.0 (pH = 10.0) 60.5 86.3 29.171.2 Tap (pH = 4.0) 68.2 90.2 32.0 66.1 (pH = 7.0) 72.1 85.2 23.5 65.9(pH = 10.0) 76.7 94.1 37.4 64.6

As Table 1 shows, including a dispersant greatly improved spinosadconcentrations in aqueous formulations at pH levels of 4, 7, and 10 inboth soft (deionized) and hard (tap) waters. The dispersant also aidedthe resuspension properties (remixing properties) of the formulationafter quiescent settling of the solids from suspension.

EXAMPLE 2 Effectiveness of Aqueous Suspension Formulation of Spinosad (1g/L) as an Ectoparasiticide

A dip formulation study for the control of Bovicola ovis Hartley Strainon sheep was conducted. In this study an aqueous suspension (AS)formulation containing 1 g/L of spinosad was prepared and diluted 1:5000in water to form a dip solution with a spinosyn concentration of 0.2ppm. Duration of study was 56 days, with lice counts taken initially, 7,14, 28, 42, and 56 days after treatment.

Table 2 summarizes the results of this study.

TABLE 2 Lice Control in Sheep with Spinosad AS Dip Formulation GroupLice Counts (geometric mean) Days post-treatment 0 7 14 28 42 56 Control203.4 187.3 180.1 219.8 208.5 199.1 Aq Suspension (0.2 ppm) 188.5 6.55.8 11.4 18.5 18.3As Table 2 shows, the dip containing 0.2 ppm of spinosad gave excellentlice control on sheep initially and after 56 days. A preferred dose ofspinosad in dip water for 100% effective lice control is considered tobe 1.0 ppm, to allow for a 5-fold confidence factor.

EXAMPLE 3 Spinosad Formulation Stability Studies

Study 1

Several spinosad-containing dip formulations were subjected to prolongedstorage at 40° C. in a chemical storage stability study. Dips 1 and 2were emulsifiable concentrates of spinosad. In Dip 1 the spinosad wasformulated in an aromatic hydrocarbon solvent with a specific gravity of0.9 at 60 F (Aromatic 150), and in Dip 2 it was formulated in methyloleate as the solvent. Dip 3 was an aqueous suspension of spinosad. Theconcentration of spinosad present in the formulations was measured afterthe compositions were exposed to this elevated temperature for 0, 7, 14,28 and 87 days. Measurements were made by analytical HPLC. Initial (Day0) measurements were listed as 100% for purposes of comparing theconcentrations of active ingredient present at later times. The resultsof this study are listed in Table 3.

TABLE 3 Spinosad Stability in Three Dip Formulations at 40° C. SpinosadConcentration, Percent of Initial Dip 1 Dip 2 Dip 3 ConcentrationsConcentrations Concentrations 0.2 g/L 1 g/L 0.2 g/L 1 g/L 0.2 g/L 1 g/LInitial Day 100 100 100 100 100 100 Day 7 114.2 86.2 91.3 92.9 105.8103.6 Day 14 95.2 76.7 86.9 89.7 100 95.1 Day 28 80.9 59.4 85.6 85 105.8100 Day 87 19 43.1 82.6 110.2 111.7 97.5

As Table 3 shows, the emulsifiable concentrate formulations were notstable, but the aqueous suspension formulation was chemically stable.

Study 2:

Two formulations containing 25 g/L of spinosad were prepared, one anaqueous suspension (AS) and the other an aqueous solution. They werestored at both ambient temperature (i.e., room temperature) and at 50°C. to compare the stability of the two formulations. HPLC quantitationwas used to measure spinosad concentration at various points in time.The results of this study are listed in Table 4.

TABLE 4 Stability of Spinosad at 25 g/L in Aqueous Suspension andSolution Formulations Spinosad Concentration, Percent of Initial AqueousSuspension Aqueous Solution Days Ambient 50° C. Ambient 50° C. 0 100 100100 100 7 107.7 100 100.3 93.4 14 115.4 107.7 98.6 88.9 21 102.3 119.299.3 77.9 28 95.7 97.7 96.9 73.4 42 97.7 98.5 96.7 67.1 70 101.5 105.479.9 50.8 98 103.8 104.2 56.9 —

At this study shows, the spinosad 25 g/L aqueous suspension waschemically stable at temperatures ranging from ambient to approximately50° C. for 98 days; however, the spinosad aqueous solutions did notexhibit long-term chemical stability at either ambient or elevatedtemperatures.

EXAMPLE 4 Spinosad Stripping from Dip Water

Two trials were conducted to determine the degree to which spinosad isremoved (stripped) from dip water after dipping sheep. In the trials,spinosad dip tank concentrations of 50 ppm and 5 ppm were prepared froman aqueous suspension containing 25 g/L of spinosad. Both trialsinvolved dipping 10 Dorset-cross shorn sheep in 70 gallons (265 liters)of treated water. Each animal was dipped for 30 seconds duration, withthe head immersed twice. Samples of dip water were taken for HPLCanalysis, initially and after each sheep was dipped. The pH of the waterwas also determined after each animal was dipped. A dip tankconcentration of 5 ppm of spinosad should provide lice control on sheep.Spinosad is in true solution at 5 and 50 ppm concentration.

During the 50 ppm trial, approximately 1.5 gallons (5.7 liters) of dipwater were lost from the dip tank for each animal dipped. The pH of thedip water increased after each sheep was dipped in the tank. Theconcentration of spinosad in the dip water decreased approximately 4%after dipping ten sheep.

During the 5 ppm trial, approximately 2.1 gallons (8 liters) of dipwater were lost from the dip tank for each animal dipped. Again, the pHof the dip water increased after each sheep was dipped in the tank. Theconcentration of spinosad in the dip water decreased approximately 14%after dipping ten sheep.

Thus, the study showed similar changes in pH and active ingredientconcentration associated with dipping sheep in diluted aqueoussuspensions with final concentrations of 50 ppm and 5 ppm. These sheepstudies indicated minimal stripping of spinosad from the diluted dipwater after dipping a limited number of animals.

EXAMPLE 5 Aqueous Suspension Formulation of Spinosad (25 g/L) Preparedby Batch Wet Milling Process

A 25 g/L concentrate of spinosad having the following components wasprepared as follows:

Component Quantity, % w/w Spinosad, technical grade @ 92.0% 2.7Dispersant Solution (LOMAR PWA) (44%) 4.5 Mineral Thickener (VEEGUM) 1.0Antimicrobial Agent (PROXEL GXL) 0.2 Propylene Glycol 10.0 Xanthan Gum(RHODOPOL 23) 0.2 Surfactant (PLURONIC P123) 1.0 Antifoam, 30% solution(ANTIFOAM C) 0.2 Deionized Water 80.2

Three stock solutions were prepared in separate agitated stainless steelvessels. A 10% stock solution (Solution A) of the surfactant was made.The surfactant used was a paste at 20° C. and so was warmed to 50° C. toliquefy it. Moderate mixing was required for its dissolution/dispersionin water.

A second stock solution (Solution B) of the mineral thickener as a 5-10%hydrate (with 5% being most typical) was prepared in water using a highshear mixer (i.e. Cowles disperser) to assure dissolution/dispersion.The cycle time required was approximately 4 hours.

In the third stock solution (Solution C), the xanthan gum was hydratedby making a slurry of the dried powder in propylene glycol containingthe full amount of the antimicrobial agent and dispersing the slurry inwater under moderate shear. Typically, xanthan gum hydrates are preparedat the 1-2% level by weight, with 1.5% being preferred. All thepropylene glycol may be used at this stage, or some may be added to thepre-mill vessel if desired.

Stock Solution A, the dispersant, deionized water, and the antifoam wereadded to a pre-mill vessel. Any propylene glycol not used in thepreparation of Solution C can be added here as well. The contents weremixed until homogeneous. The full amount of Solution B can be added atthis stage or delayed until recovery of the post-mill material.

Next, the spinosad was added slowly under moderate agitation. After theaddition was complete, it was necessary to increase shear to preventfloating and assure wetting. To assure appropriate wetting and reductionof spinosad clumps, the use of a stainless steel rotor/statorhomogenizer is recommended, with its effluent recirculated back into thepre-mill vessel.

The contents of the pre-mill vessel were displaced to a wet milloperation (stainless steel bead mill, horizontal preferred) to achievefurther particle size reduction. The milled material and the millrinsate were collected in an agitated, tared, stainless steel vessel(post-mill vessel). The amount of post-mill material recovered wasrecorded and compared relative to theoretical recovery to determine thepercent recovery.

From the percent recovery, the exact amount of Solution C (hydratedxanthan gum) necessary to provide the final product was calculated. IfSolution B was not added at the pre-mill stage, use the same calculationperformed with Solution C to determine the amount of Solution B to addnow. The calculated amount of Solution C (and Solution B if necessary)was added to the post-mill material with mild agitation (propeller bladeat roughly 500 rpm), and allowed to stir for 1 hour. A sample of finalproduct was taken to assay for viscosity, particle size and specificgravity.

EXAMPLE 6 Aqueous Suspension Formulation of Spinosad (0.2 g/L)

An aqueous suspension containing 0.2 g of spinosad/L was prepared asfollows:

Quantity Component % w/w Batch (g) Spinosad, technical, 92.1% 0.02 0.03Dispersant 0.1 0.15 Propylene Glycol 10 15 Surfactant 2 3 MineralThickener 2 3 Antimicrobial Agent 0.2 0.3 Xanthan Gum 0.2 0.3 Water,deionized 85.38 128.07 Antifoam 0.1 0.15 100 150

To prepare the 0.2 g/L aqueous suspension, the following steps aretaken: spinosad technical grade (0.03 g) is mixed with surfactant (3.0g), dispersant (0.15 g), deionized water (7.52 g) and antifoam (0.15 g)and wet-milled in a ball mill to form a grind batch.

Separately, a hydrate suspension batch is prepared as follows: addxanthan gum (0.3 g) to propylene glycol (15 g) and antimicrobial agent(0.3 g) with mixing. Add mineral thickener (3 g) to deionized water(120.55 g) with mixing. Allow the xanthan gum to fully disperse withinthe propylene glycol; then add the xanthan gum/propylene glycol to thehydrated mineral thickener with mixing.

To form the final aqueous suspension product, add the grind batch to aquantity of the hydrate solution sufficient to form an aqueoussuspension with a final spinosad concentration of 0.2 g/L.

EXAMPLE 7 Aqueous Suspension Pour-On Formulation of Spinosad (100 g/L)Containing a Substantive Agent

An aqueous suspension formulation containing 100 g/L of spinosad and apolyvinyl acetate is prepared. The polyvinyl acetate acts as asticker/binder component to increase the formulation's adhesion to hairwhen used as a pour-on. The suspension components are:

Quantity Component % w/w Batch (g) Spinosad, technical 11.04 5.52Dispersant, 44% solution 8 4 Propylene Glycol 20 10 Surfactant 1 0.5Mineral Thickener 2 1 Polyvinyl Acetate 10 5 Water, deionized 47.6623.83 Antimicrobial Agent 0.2 0.1 Antifoam 0.1 0.05 100 50

For this formulation, the grind batch is produced by wet-milling thespinosad in 10 g of deionized water containing the surfactant, thedispersant solution, and 0.05 g of the antifoam. The grind batch isrinsed with an additional 5 g of water. Recovery of the grind batch is90.46%.

The hydrated suspension is formed by mixing the mineral thickener into8.83 grams of water containing the antimicrobial agent. The polyvinylacetate is added to the propylene glycol. The mineral thickener hydrateand the polyvinyl acetate/propylene glycol mixture are mixed to form thehydrate suspension.

The hydrated suspension is added to the grind batch and mixed untiluniform.

EXAMPLE 8 Aqueous Suspension Pour-on Formulation of Spinosad (100 g/L)

A pour-on formulation containing 100 g/L of spinosad is prepared asfollows:

Quantity Component % w/w Batch (g) Spinosad, technical 11.04 5.52Dispersant, 44% solution 8 4 Propylene Glycol 10 5 Surfactant 1 0.5Mineral Thickener 2 1 Polyvinyl Alcohol, 10% 20 10 Solution Water,deionized 47.66 23.83 Antimicrobial Agent 0.2 0.1 Antifoam 0.1 0.02 10050

The grind batch is formed by wet-milling the spinosad in 10 g ofdeionized water containing the surfactant, the dispersant solution, andthe antifoam. The grind batch is rinsed with an additional 5 g of water.Recovery of the grind batch is 88.56%.

The 10% (w/w) solution of polyvinyl alcohol (AIRVOL 125™, Air Products)is formed by mixing it in deionized water heated to 96° C. until it isin solution and allowing the solution to cool to room temperature.

The hydrated suspension batch is formed by mixing the propylene glycol,the mineral thickener, the antimicrobial agent, the polyvinyl alcoholand deionized water (8.83 grams).

The final product is formed by adding 88.56 percent of the hydratedsuspension to the grind batch and mixing until uniform.

EXAMPLE 9 Aqueous Suspension Formulation of Spinosad (25 g/L) withPolymeric Thickener and Polymeric Dispersant

An aqueous suspension of spinosad was prepared as follows:

Quantity Component % w/w Batch (g) Spinosad, technical (92%) 2.6 26Antimicrobial Agent 0.25 2.5 Antifoam 0.1 1 Surfactant 2 20 PolymericDispersant 3 30 Polymeric Thickener 4 40 Propylene Glycol 6 60 Water,deionized 82.05 820.5 100 1000

The grind batch was formed by wet-milling the spinosad in 64.9 g ofdeionized water containing the antimicrobial agent, 0.1 g of theantifoam, 2 g of the surfactant, 2 g of the polymeric dispersant and 2.5g of the polymeric thickener. Recovery of the grind batch was 91%. Atypical polymeric surfactant is ATLOX 4894, and a useful polymericdispersant is ATLOX 4913 (both manufactured by Uniqema).

A letdown batch was separately formed by siring together 0.9 g of theantifoam, 18 g of the surfactant, 28 g of the polymeric dispersant, 37.5g of the polymeric thickener, the propylene glycol and 755.6 g ofdeionized water. The final product was formed by adding 819 g of theletdown batch to the grind batch and mixing until uniform.

EXAMPLE 10 Aqueous Suspension Formulation of Spinosad (480 g/L)

An aqueous suspension containing 480 g/L of spinosad is prepared asfollows:

Quantity Component % w/w Batch (g) Spinosad, technical (91.4%) 49.14196.56 Propylene Glycol 3 12 Surfactant 3 12 Lignosulfonate Dispersant14 56 Xanthan Gum 0.05 0.2 Mineral Thickener 0.35 1.4 Water, deionized29.76 119.04 Antimicrobial Agent 0.2 0.8 Antifoam 0.5 2 100 400

Mix the spinosad with the surfactant, the dispersant, deionized water(90.74 g) and 2 g of antifoam, and wet-mill in a ball mill to form agrind batch. A suitable lignosulfonate dispersant is Reax 88B, WestvacoCorporation, Inc. Recovery of the grind batch was 66.20%.

Separately, prepare the hydrate suspension batch as follows: add thexanthan gum to the propylene glycol with mixing, then add the mineralthickener to 28.3 g of deionized water and the antimicrobial agent withmixing. Allow the xanthan gum to fully disperse within the propyleneglycol before adding the xanthan gum/propylene glycol to the hydratedmineral thickener with mixing.

The final product is formed by adding 28.27 grams of the hydratedsuspension to the grind batch and mixing until uniform.

EXAMPLE 11 Aqueous Suspension Pour-On Formulation of Spinosad (100 g/L)with Sucrose

The following components are used in this formulation:

Quantity Component % w/w Batch (g) Spinosad, technical (90%) 9.55 28.65Propylene Glycol 5 15 Surfactant 1.5 4.5 Dispersant, 44% Solution 8 24Sucrose Solution, 40% 75.55 226.65 Antimicrobial Agent 0.2 0.6 Antifoam0.2 0.6 100 300

To form the grind batch, combine the spinosad, the surfactant, thedispersant, the antifoam, and the 40% aqueous sucrose solution and wetmill to desired spinosad median particle size. Recover milled materialfrom mill and determine the percent recovery. To the recovered grindmaterial, add the appropriate amount of propylene glycol andantimicrobial agent to bring the final concentration of spinosad to 100g/L.

EXAMPLE 12 Aqueous Suspension Pour-On Formulation of Spinosad (200 g/L)with Urea

The following components are used in this formulation:

Quantity Component % w/w Batch (g) Spinosad, technical (90%) 19.7 59.1Propylene Glycol 5 15 Surfactant 2 6 Dispersant, 44% Solution 14 42Urea, aqueous solution, 50% 58.9 176.7 Antimicrobial Agent 0.2 0.6Antifoam 0.2 0.6 100 300

To form the grind batch, combine the spinosad, the surfactant, thedispersant, the antifoam, and the urea solution and wet mill to desiredspinosad median particle size. Recover milled material from mill anddetermine the percent recovery. To the recovered grind material, add theappropriate amount of propylene glycol and antimicrobial agent to bringthe final concentration of spinosad to 200 g/L.

1. A stable ectoparasiticidal aqueous suspension formulation suitablefor administration to animals comprising: a) an ectoparasiticidal amountof spinosad, milled to an average particle size of from 1 to 15 microns;b) a condensed formaldehyde/naphthalene sulfonic acid or salt thereofdispersant, wherein the spinosad:dispersant weight ratio is 3:1 to 1:5;c) 0.3 to 5 weight percent of a mineral thickener; d) an antimicrobialagent acceptable for topical veterinary applications in an amounteffective to prevent microbial growth in the suspension; e) propyleneglycol; f) 0.05 to 3 weight percent of xanthan gum; g) 0.1 to 10 weightpercent of at least one surfactant; h) an antifoam agent; and i) water;wherein said formulation is chemically and physically stable.
 2. Aformulation of claim 1 wherein the average particle size of spinosad isabout 2 to about 7 microns.
 3. A formulation of claim 2 wherein theamount of spinosad is from about 0.02 to about 50 weight percent of theformulation.
 4. A formulation of claim 3 wherein the amount of spinosadis from about 2 to about 5 weight percent of the formulation.
 5. Aformulation of claim 4 wherein the surfactant is present in an amount offrom about 0.1 to about 5 weight percent of the formulation.
 6. Aformulation of claim 5 wherein the spinosad is present in an amount ofabout 25 grams per liter of the formulation.
 7. An article ofmanufacture, comprising packaging material and a formulation forcontrolling an ectoparasite infestation on a small ruminant or companionanimal contained within said packaging material, wherein saidformulation comprises: a unit dose of a formulation of claim 1; andwherein said packaging material comprises a label or package insert withinstructions for administering the dose to the animal.
 8. A method ofcontrolling an ectoparasite infestation on a small ruminant or companionanimal, comprising administering to the animal an effective amount offormulation of claim
 1. 9. The method of claim 8 wherein the formulationis applied to the head, neck, shoulders or back of the animal by aspot-on or pour-on protocol.